Abstract
The salivary gland is a key organ in insects that plays essential roles in food digestion, nutrient absorption, and energy metabolism, thereby highlighting the importance of studying salivary gland function for gaining a better understanding of nutritional utilization and insect-plant interactions. To date, however, a lack of salivary gland-specific promoters has limited functional analyses of salivary gland genes in Lepidoptera. In this study, based on microarray and salivary gland transcriptome data, we identified nine candidate genes characterized by high salivary gland expression. Semi-quantitative PCR analysis confirmed cholinesterase (BmCe, BGIBMGA010988) as the optimal candidate for promoter cloning. Temporal expression analysis revealed that the expression of BmCe reaches a peak during days 2-4 of the fifth larval instar. A 2152 bp fragment upstream of the transcription initiation site of BmCe was selected as the putative promoter sequence (designated BmCeP) and cloned to construct a piggyBac transgenic vector driving DsRed expression. Transgenic silkworms were obtained via embryonic microinjection and tissue expression analysis on day three of fifth-instar larvae revealed the predominant localization of DsRed expression in the salivary glands. In this study, we thus identified a gene promoter characterized by salivary gland-predominant expression in Bombyx mori, which we believe could serve as a valuable genetic tool for investigating the molecular mechanisms underlying silkworm nutritional utilization and interactions with its host plant, mulberry.